Research news

Small, noncoding RNA molecules mediate a posttranscriptional gene-silencing mechanism that regulates the expression of developmental genes by inhibiting the translation
of target mRNAs. This mechanism is common to plants, fungi, and animals, and the generation
of these microRNAs (miRNAs, also known as small inhibitory RNAs or siRNAs) involves
a series of sequential steps, where primary RNA transcripts (pri-miRNAs) are cleaved
in the nucleus to smaller pre-miRNAs. These are transported to the cytosol where Dicer,
a member of the RNAse III nuclease family, further processes them to yield mature
miRNAs. MiRNAs associate with multicomponent ribonucleoprotein complexes, or RISCs,
which effect the silencing of the target mRNA molecules. Two papers in the September
25 Nature report previously unidentified key components in the miRNA regulatory system. Yountae
Lee and colleagues at Seoul National University identify a protein that cleaves pri-miRNAs to pre-miRNAs, while Amy Caudy and colleagues
at Cold Spring Harbor Laboratory identify another component of the RISC complex.

Lee et al. cloned and sequenced the sites of cleavage of the miRNA miR-30a, revealing a stem-loop
structure with a two-nucleotide overhang suggestive of the mechanism of action of
another RNAse III family member. The authors then focused on the candidate protein
Drosha, because of its nuclear location and role in preribosomal RNA processing. Immunopurified
Drosha cleaved pri-miRNAs in vitro, while its knockout (by Drosha-specific miRNA interference)
resulted in the accumulation of pri-miRNAs and a depletion of the mature product.
Examining the action of Drosha with Dicer revealed a synergistic role of these two
nucleases in the accurate generation of specific miRNAs (Nature, DOI:10.1038/nature01957, September 25, 2003).

"Our study introduces a new player in miRNA biogenesis. It is an open question how
Drosha and Dicer recognize their targets in a specific manner when the primary sequences
of diverse miRNAs show no conserved elements. In view of this, it will be important
to determine structures of miRNA precursors and RNase III proteins," conclude Lee
and colleagues.

Caudy et al. purified a RISC complex from Drosophila and identified its protein and RNA components. They identified an evolutionarily
conserved 103-kDa protein containing five repeats of a staphylococcal/micrococcal
nuclease domain, with the fifth domain fused to a tudor domain - hence the name Tudor-SN. The authors also identified this molecule in RISCs
from Caenorhabditis elegans and mammals with similar functional activity to other staphylococcal nucleases and
showed that a specific competitive micrococcal nuclease inhibitor inhibited both Tudor-SN
and RISCs (Nature, DOI:10.1038/nature01956, September 25, 2003).

"Our data strongly indicate that Tudor-SN is a bona-fide RISC component. It is certainly
consistent with our biochemical data to suppose that RISC contains multiple nucleases,
only one of which... can catalyze site-specific mRNA cleavage. In this scenario, Tudor-SN
might act to degrade the remainder of the mRNA," conclude Caudy and colleagues.